Our overall aim is to identify and characterize basic mechanisms underlying defective growth control of cancer cells. Much evidence now supports our hypothesis that normal cell proliferation is controlled just prior to the sudden onset of DNA synthesis and related biochemical processes. We have been able to apply molecular biology techniques by developing a productive "reverse biochemistry" approach which starts with the appearance of mouse TK mRNA as a terminal marker of the Gl/S transition. Our most recent work has implicated Gl/S Yi complexes that contain a DNA binding protein (p6O), a cdc2 kinase, a retinoblastoma-like 110 KD phosphoprotein (pRBm), and a cycle limiting factor that may be a cyclin. We are bringing together these molecules and the cell biology of proliferation control. (I) Identify and characterize the Yi protein complexes that bind to consensus promoter sequences in murine TK. Clone genes to obtain probes and antibodies related to Yi proteins. (II) Investigate the relevance of this Yi system in deranged control of cells from tumors and by viral transformation. We will particularly determine the molecular change that confers stability to the Yi complex in tumor cells. (III) Investigate mechanisms leading to assembly of cyclins, CDC2 kinase and phosphorylation of pRB, which activate binding of Yi to its DNA consensus sequence, and conversion of the Gl-form to the S-form of Yi. (IV) Generalize our observations regarding the Yi binding system and its preceding regulatory process to other G1/S transcriptions and cell proliferation control. Computer data banks will be searched for genes whose promoters contain similar consensus sequences. Initially, a study in depth will be made of the clinically important, cycle regulated enzyme thymidylate synthase (TS). We will use human breast cells because of their relevance, suitability and availability to studies of cancer therapy, prognosis and diagnosis.